IDENTIFICATION AND EXPRESSION ANALYSIS OF ABC GENES IN PLASMODIUM YOELII

约氏疟原虫ABC基因的鉴定及表达分析

• 批准号: 7610152
• 负责人: IVAN FERRER
• 金额: $ 25.65万
• 依托单位: UNIVERSITY OF PUERTO RICO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7610152
• 关键词: Address; Anopheles gambiae; Anti-malarial drug resistance; Antimalarials; Binding; Bioinformatics; Carrier Proteins; Collection; Computer Retrieval of Information on Scientific Projects Database; Disruption; Drug Design; Drug resistance; Drug-sensitive; Evolution; Facility Construction Funding Category; Funding; Gene Dosage; Gene Proteins; Genes; Genome; Goals; Grant; Health; Homologous Gene; Institution; Lead; Location; Malaria; Measures; Mediating; Membrane; Modeling; Molecular; Multi-Drug Resistance; Multidrug Resistance-Associated Proteins; Numbers; Orthologous Gene; Parasites; Parasitic Diseases; Pharmaceutical Preparations; Plasmodium; Plasmodium yoelii; Proteins; Recombinant Proteins; Research; Research Personnel; Resistance; Resources; Rodent; Role; Single Nucleotide Polymorphism; Source; Transcript; Transfection; United States National Institutes of Health; Work; base; disorder control; in vivo; innovation; member; novel; quinoline; resistance mechanism

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Malaria is one of the most important parasitic diseases worldwide. The evolution and spread of multidrug resistant strains of P. falciparum and P. vivax is a major health concern. Despite the advances in research, including the sequencing of the entire genomes of Anopheles gambiae, P. falciparum and P. yoelii, the molecular basis of the drug resistance phenomena in Plasmodia is not completely understood. The long-term goal of the proposed research is to elucidate mechanisms by which the malaria parasites become resistant to antimalarial drugs. The results of this research could lead to a better approach to drug design and disease control. Our working hypothesis is that several genes that belong to the ATP-binding cassette (ABC) superfamily of transporter proteins work in conjunction to confer drug resistance. A collection of drug sensitive and resistant lines of P. yoelii will be used as an in vivo rodent malaria model to investigate the mechanisms of drug resistance. Although a number of resistance mechanisms are possible, we will focus on two members of the ABC superfamily of transporter proteins that may mediate drug efftux in the parasite, the multidrug resistance associated protein gene (pymrp) and the ABCG homologue gene. The specific aims are: - To characterize the P. yoelii multidrug resistance associated protein gene (pymrp) and to determine its role in malaria drug resistance. - To measure transcript levels of the pymrp gene in drug sensitive and resistant lines. - To determine the sub-cellular location and measure expression levels of the pyMRP protein. - To initiate transfection studies to address the function of the pyMRP protein by gene disruption. - To characterize a novel ABC gene, the P. yoelii ABCG homologue, and determine its role in malaria drug resistance. - To determine membrane topology and identify conserved motifs in the P. yoelii ABCG homologue by performing bioinformatic analyses. - To analyze the P. yoelii ABCG homologue, from drug sensitive and resistant lines, for gene copy number, single nucleotide polymorphisms (SNP), transcript levels and chromosomal location. - To determine the sub-cellular location and measure expression levels of the P. yoelii ABCG protein. The rational for this research is that understanding the role of pymrp and the P. yoelii ABCG homologue gene in Plasmodium drug resistance could provide the basis for better drug design and potentially lead to reversal of drug resistance to the quinoline-containing antimalarial drugs. This is an innovative research because the function of the Plasmodium mrp and ABCG ortholog genes and their potential role in drug resistance remains to be elucidated. During the first year we expect to accomplish the following goals: 1) To measure transcript levels of the pymrp gene in drug sensitive and resistant lines; 2) To determine membrane topology and identify conserved motifs in the P. yoelii ABCG homologue by performing bioinformatic analyses; and 3) To initiate the construction of the pyMRP recombinant protein.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 疟疾是全世界最重要的寄生虫病之一。恶性疟原虫和间日疟原虫多重耐药菌株的进化和传播是一个主要的健康问题。尽管研究取得了进展，包括对冈比亚按蚊、恶性疟原虫和约氏疟原虫的整个基因组进行测序，但疟原虫耐药现象的分子基础尚未完全了解。拟议研究的长期目标是阐明疟原虫对抗疟药物产生耐药性的机制。这项研究的结果可能会带来更好的药物设计和疾病控制方法。我们的工作假设是，属于转运蛋白 ATP 结合盒 (ABC) 超家族的几个基因协同作用，产生耐药性。约氏疟原虫对药物敏感和耐药的品系集合将被用作体内啮齿动物疟疾模型，以研究耐药机制。尽管可能存在多种耐药机制，但我们将重点关注转运蛋白 ABC 超家族的两个成员，它们可能介导寄生虫中的药物流出，即多药耐药性相关蛋白基因 (pymrp) 和 ABCG 同源基因。 具体目标是： - 表征约氏疟原虫多药耐药性相关蛋白基因 (pymrp) 并确定其在疟疾耐药性中的作用。 - 测量药物敏感和耐药品系中 pymrp 基因的转录水平。 - 确定 pyMRP 蛋白的亚细胞位置并测量表达水平。 - 启动转染研究，通过基因破坏来解决 pyMRP 蛋白的功能。 - 表征新型 ABC 基因（约氏疟原虫 ABCG 同源物），并确定其在疟疾耐药性中的作用。 - 通过进行生物信息学分析来确定膜拓扑结构并鉴定 P. yoelii ABCG 同源物中的保守基序。 - 分析来自药物敏感和耐药品系的 P. yoelii ABCG 同源物的基因拷贝数、单核苷酸多态性 (SNP)、转录水平和染色体位置。 - 确定约氏疟原虫 ABCG 蛋白的亚细胞位置并测量表达水平。 这项研究的理由是，了解 pymrp 和约氏疟原虫 ABCG 同源基因在疟原虫耐药性中的作用可以为更好的药物设计提供基础，并有可能逆转含喹啉抗疟药物的耐药性。这是一项创新性研究，因为疟原虫 mrp 和 ABCG 直系同源基因的功能及其在耐药性中的潜在作用仍有待阐明。在第一年中，我们期望实现以下目标： 1) 测量药物敏感和耐药品系中 pymrp 基因的转录水平； 2) 通过生物信息学分析确定膜拓扑结构并鉴定 P. yoelii ABCG 同源物中的保守基序； 3) 启动pyMRP重组蛋白的构建。